Introduction and Background of the Project

Introduction

A large proportion of people living with HIV die from undiagnosed TB, due to the absence of appropriate diagnosis and treatment. Implementation of systematic screening for TB among high-risk groups is expected to be a key intervention to reduce TB deaths by 75% by 2025. Achieving this ambitious goal will require novel, sensitive point-of-care tests for the detection of active TB in HIV-positives. Existing tests cannot detect TB among a substantial proportion of patients with HIV in decentralized settings due to the size and complexity of the required instrumentation and because a substantial proportion of patients with HIV-associated TB cannot produce sputum. Therefore a rapid, sensitive, biomarker-based, non-sputum-based point-of-care diagnostic that makes use of an easily accessible sample like blood or urine and is able to diagnose TB in HIV-infected individuals is considered to be one of the most urgently needed tests by expert stakeholder groups.

Project objective

Develop a highly sensitive, rapid point-of-care TB test to diagnose TB from urine in high-risk groups. Such a test will enable early treatment initiation, decrease mortality and cut transmission.

Project design

FIND and FUJIFILM’s project aims to: 1) Validate and select reagents to be used in the development of a LAM-based diagnostic test Lipoarabinomannan (LAM) is an antigen that has been explored extensively in blood, urine and sputum as a biomarker for active TB detection. It has the advantage of being very stable and detectable in a wide diagnostic time window. 2) Based on the selected reagents, develop a sensitive, point-of-care, rapid diagnostic test for the detection of TB infection in HIV infected patients, based on Fujifilm technology 3) Collect well-characterized patient samples from tuberculosis patients with and without HIV-coinfection, including endemic controls, and use the collected specimens to conduct an independent evaluation of the test under development.

How can your partnership (project) address global health challenges?

TB is the most common cause of mortality in HIV patients. TB/HIV-coinfection often goes undiagnosed: first, many patients are unable to produce sputum (necessary for current diagnostics) and symptoms of TB in HIV patients vary greatly. Therefore, a test that uses either urine or blood (instead of sputum) which can be done at the point-of-care to screen HIV patients is urgently needed.

What sort of innovation are you bringing in your project?

Currently, no simple, rapid and affordable POC TB test with sufficient performance is available. In a priority-setting exercise involving different groups of global stakeholders (including national TB programmes), the non-sputum-based biomarker test was rated highest by all stakeholder groups. This project is a major step towards a test for the systematic screening of TB in the high-risk group of HIV-positives. The proposed test is expected to show increased sensitivity in HIV-positive patients, which would significantly decrease morbidity and mortality. If the developed test achieves sensitivity and specificity high enough to work as a general rapid biomarker-based POC test for TB (including HIV-negative adults and children), it would be a real game-changer for TB diagnosis. A modeling has demonstrated that a 16% decrease in TB incidence and a 39% decrease in mortality(1) if our project succeeds.

Role and Responsibility of Each Partner

Fujifilm will be responsible for development, optimization and manufacturing of the assay prototype, as well as the provision of sufficient numbers of tests for the independent evaluation. FIND will be the overall coordinator of the project, responsible for establishing project goals and project management, monitoring and reporting on progress, collecting and providing reference materials, and engaging additional partners if needed. FIND will supply reagents and samples, set the product specifications, establish the reference assay and organize the independent evaluation with a WHO collaborating Centre for TB and Lung Diseases and conduct the statistical analysis of the obtained data.

Final Report

1. Project objective

Introduction and Background of the Project:

Tuberculosis (TB) is the number one infectious disease killer globally. A large proportion of people living with HIV (PLHIV) die from undiagnosed TB due to the absence of appropriate diagnosis and treatment. The End TB strategy aims for a 90% reduction in TB morality and an 80% reduction in TB infection by 2030. Achieving this goal requires novel, sensitive point-of-care (POC) tests for detecting active TB in PLHIV.

Project objective

Fujifilm and FIND have been working together since 2015 to develop a sensitive, rapid POC TB test to diagnose TB from urine. The targeted test should enable early treatment initiation, decrease mortality and cut transmission.

2. Project design

As part of this project, the partnership:

(a) screened a large number of antibodies to find the best candidates for the development of a lipoarabinomannan (LAM)-based diagnostic test;

(b) used the best antibody combination to design and develop a sensitive POC rapid diagnostic test based on Fujifilm’s innovative silver amplification technology;

(c) collected urine samples from tuberculosis patients, and used the collected specimens to conduct a study to assess the performance of the novel test.

3. Results, lessons learned

Fujifilm and FIND finalized the design and development of the POC test to diagnose TB in PLHIV, completing 3 GHIT stage-gates and achieving 9 milestones.

(a) Screening for the best antibody pair

First, FIND and partners evaluated >1000 monoclonal antibody pairs on a sensitive laboratory platform to find the most suitable pair for the POC test. Surprisingly, many pairs showed high reactivity to purified LAM, but performed poorly at detecting urinary LAM in clinical samples, suggesting differences in antigen structure and immunoreactivity of the different LAM sources. Therefore it was critical to use TB patient urine for the screening. The best antibody pair reached femtomolar analytical sensitivity for LAM detection and a high clinical sensitivity and specificity.1

(b) Design and development of FUJIFILM SILVAMP TB LAM

Next was to convert the complex, 2-hour laboratory platform assay run on an expensive and sophisticated analyzer into a simple, fully-disposable and easy-to-use POC test. Fujifilm used its silver amplification technology2 that works like a photographic development step for the first time in a urine-based test and also designed and developed a manual cartridge that does not require an instrument, thus meeting the needs of low-resource settings. The Fujifilm R&D team did several rounds of assay optimization to enhance the sensitivity and shorten the assay time. The resulting FUJIFILM SILVAMP TB LAM (FujiLAM) is probably the most sensitive POC lateral flow immunoassay in the world and is able to measure very low concentrations of LAM in urine.

(c) Evaluating diagnostic accuracy

As a third step, FIND performed an independent clinical study to evaluate the diagnostic accuracy of FujiLAM on banked samples in South Africa. Nearly 1000 PLHIV were evaluated and FujiLAM detected 28% more TB patients compared to the only currently available POC test. In conclusion, FujiLAM offers superior diagnostic sensitivity, while maintaining specificity and has the potential to transform rapid POC TB diagnosis for PLHIV. 3

Next steps are to launch FujiLAM as an IVD product and organize large scale prospective trials to inform WHO policy with the goal to use the test for TB diagnosis in PLHIV in LMICs.

3Broger, T et al. Novel high sensitivity tuberculosis point-of-care test for people living with HIV. Under review. Available at https://ssrn.com/abstract=3254479 (Comment: this manuscript is currently under peer-review and on a pre-print server. We recommend to wait with linking this paper until it has been accepted by the journal)